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P2072
Label-Free Determination of Kinetic Constants for Small Molecule Binding to Proteins Using FortéBio’s Octet RED Multi-Channel System
Presenter Charles Wartchow, ForteBio, USA
Additional Authors: Danfeng Yao, Pu Li, Bettina Heidecker, Jing Wei, Sae Choo, Robert Zuk
The determination and evaluation of the affinity of small molecule binding to a therapeutic target is a significant component of drug discovery and lead optimization.  We report a label-free method for determining kinetic constants for the binding of small molecule inhibitors (<500 Daltons) to carbonic anhydrase, including rate constants and affinity constants.  These parameters are derived from data obtained using FortéBio’s Octet RED System, a label-free, 8-channel system that is compatible with 96-well microplates.    The system is based on BioLayer Interferometry, a technique that generates an interference pattern by monitoring visible light reflected from two surfaces in a fiber-optic biosensor.  When binding events occur at the tip of a fiber optic biosensor, the interference pattern shifts to higher wavelength, and these changes are monitored in real time.  In a typical experiment, biotinylated protein targets are immobilized onto a high-binding capacity streptavidin surface, and the association and dissociation of small molecules with molecular weights ranging from ~150-350 Daltons is monitored in parallel.  Kinetic constants for the binding of compounds to carbonic anhydrase with micromolar and nanomolar affinities agree closely with published values.  In addition, evaluations conducted on proteins obtained from major pharmaceutical partners show agreement of the results between Octet RED and other SPR-based methods.